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6'-?-Fluoro-Homoaristeromycin and 6'-Fluoro-Homoneplanocin A Are Potent Inhibitors of Chikungunya Virus Replication through Their Direct Effect on Viral Nonstructural Protein 1.

ABSTRACT: Alphaviruses are arthropod-borne, positive-stranded RNA viruses capable of causing severe disease with high morbidity. Chikungunya virus (CHIKV) is an alphavirus that causes a febrile illness which can progress into chronic arthralgia. The current lack of vaccines and specific treatment for CHIKV infection underscores the need to develop new therapeutic interventions. To discover new antiviral agents, we performed a compound screen in cell culture-based infection models and identified two carbocyclic adenosine analogues, 6'-?-fluoro-homoaristeromycin (FHA) and 6'-fluoro-homoneplanocin A (FHNA), that displayed potent activity against CHIKV and Semliki Forest virus (SFV) with 50% effective concentrations in the nanomolar range at nontoxic concentrations. The compounds, designed as inhibitors of the host enzyme S-adenosylhomocysteine (SAH) hydrolase, impeded postentry steps in CHIKV and SFV replication. Selection of FHNA-resistant mutants and reverse genetics studies demonstrated that the combination of mutations G230R and K299E in CHIKV nonstructural protein 1 (nsP1) conferred resistance to the compounds. Enzymatic assays with purified wild-type (wt) SFV nsP1 suggested that an oxidized (3'-keto) form, rather than FHNA itself, directly inhibited the MTase activity, while a mutant protein with the K231R and K299E substitutions was insensitive to the compound. Both wt nsP1 and the resistant mutant were equally sensitive to the inhibitory effect of SAH. Our combined data suggest that FHA and FHNA inhibit CHIKV and SFV replication by directly targeting the MTase activity of nsP1, rather than through an indirect effect on host SAH hydrolase. The high potency and selectivity of these novel alphavirus mRNA capping inhibitors warrant further preclinical investigation of these compounds.

SUBMITTER: Kovacikova K

PROVIDER: S-EPMC7179274 | biostudies-literature | 2020 Mar

REPOSITORIES: biostudies-literature

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6'-β-Fluoro-Homoaristeromycin and 6'-Fluoro-Homoneplanocin A Are Potent Inhibitors of Chikungunya Virus Replication through Their Direct Effect on Viral Nonstructural Protein 1.

Kovacikova Kristina K Morren Bas M BM Tas Ali A Albulescu Irina C IC van Rijswijk Robin R Jarhad Dnyandev B DB Shin Young Sup YS Jang Min Hwan MH Kim Gyudong G Lee Hyuk Woo HW Jeong Lak Shin LS Snijder Eric J EJ van Hemert Martijn J MJ

Antimicrobial agents and chemotherapy 20200324 4

Alphaviruses are arthropod-borne, positive-stranded RNA viruses capable of causing severe disease with high morbidity. Chikungunya virus (CHIKV) is an alphavirus that causes a febrile illness which can progress into chronic arthralgia. The current lack of vaccines and specific treatment for CHIKV infection underscores the need to develop new therapeutic interventions. To discover new antiviral agents, we performed a compound screen in cell culture-based infection models and identified two carboc ...[more]

PMID: 31964798

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Project description:BackgroundChikungunya Virus (ChikV) surprised by a massive re-emerging outbreak in Indian Ocean in 2006, reaching Europe in 2007 and exhibited exceptional severe physiopathology in infants and elderly patients. In this context, it is important to analyze the innate immune host responses triggered against ChikV. Autophagy has been shown to be an important component of the innate immune response and is involved in host defense elimination of different pathogens. However, the autophagic process was recently observed to be hijacked by virus for their own replication. Here we provide the first evidence that hallmarks of autophagy are specifically found in HEK.293 infected cells and are involved in ChikV replication.MethodsTo test the capacity of ChikV to mobilize the autophagic machinery, we performed fluorescence microscopy experiments on HEK.GFP.LC3 stable cells, and followed the LC3 distribution during the time course of ChikV infection. To confirm this, we performed electron microscopy on HEK.293 infected cells. To test the effect of ChikV-induced-autophagy on viral replication, we blocked the autophagic process, either by pharmacological (3-MA) or genetic inhibition (siRNA against the transcript of Beclin 1, an autophagic protein), and analyzed the percentage of infected cells and the viral RNA load released in the supernatant. Moreover, the effect of induction of autophagy by Rapamycin on viral replication was tested.ResultsThe increasing number of GFP-LC3 positive cells with a punctate staining together with the enhanced number of GFP-LC3 dots per cell showed that ChikV triggered an autophagic process in HEK.293 infected cells. Those results were confirmed by electron microscopy analysis since numerous membrane-bound vacuoles characteristic of autophagosomes were observed in infected cells. Moreover, we found that inhibition of autophagy, either by biochemical reagent and RNA interference, dramatically decreases ChikV replication.ConclusionsTaken together, our results suggest that autophagy may play a promoting role in ChikV replication. Investigating in details the relationship between autophagy and viral replication will greatly improve our knowledge of the pathogenesis of ChikV and provide insight for the design of candidate antiviral therapeutics.

Dataset Information

6'-?-Fluoro-Homoaristeromycin and 6'-Fluoro-Homoneplanocin A Are Potent Inhibitors of Chikungunya Virus Replication through Their Direct Effect on Viral Nonstructural Protein 1.

Publications

6'-β-Fluoro-Homoaristeromycin and 6'-Fluoro-Homoneplanocin A Are Potent Inhibitors of Chikungunya Virus Replication through Their Direct Effect on Viral Nonstructural Protein 1.

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